Zinc oxide particle preparation and methods of use

ABSTRACT

A method of using ZnO particles for the treatment of colon cancer and a method of using the particles for reducing the concentration of an organic contaminant in an aqueous solution is described. The ZnO particles are substantially spherical and may have nanopetals that provide a nanoflower morphology. The synthesis and characterization of the ZnO particles is also discussed.

BACKGROUND OF THE INVENTION Technical Field

The present invention relates to a method of using ZnO particles totreat colon cancer and a method of using ZnO nanoflowers to reduce aconcentration of an organic contaminant in an aqueous solution.

Description of the Related Art

The “background” description provided herein is for the purpose ofgenerally presenting the context of the disclosure. Work of thepresently named inventors, to the extent it is described in thisbackground section, as well as aspects of the description which may nototherwise qualify as prior art at the time of filing, are neitherexpressly or impliedly admitted as prior art against the presentinvention.

Zinc oxide (ZnO) is used for various purposes including as a whitepigment, as a catalyst, as a constituent of anti-bacterial skinprotection ointment, sunscreens, and wood varnishes. Zinc oxide is alsoknown as wide band gap semiconductor and is well suited for emissivedevices. Materials used for blocking UV radiation are required to betransparent to the visible part of the solar radiation while blockingthe harmful UV radiation and zinc oxide is considered favorable in thisregard.

However, many of the above applications use nano-scale zinc oxide, andlittle progress has been made in using zinc oxide particles on largerscales having different morphologies and increased surface areas. Thoughnumerous processes are known for the synthesis of zinc oxide particles,such processes are not efficient and do not reliably produce zinc oxideparticles with high surface areas or with attached nanostructures. Thislimitation is often a significant deterrent in exploring new uses ofzinc oxide particles. In view of the foregoing, one objective of thepresent invention is to provide a method of using ZnO particles to treatcolon cancer in a mammal and a method of using the particles to reduce aconcentration of an organic contaminant in an aqueous solution byphotocatalytic degradation and/or adsorption. The ZnO particles may bemade by a specific synthesis route.

BRIEF SUMMARY OF THE INVENTION

According to a first aspect, the present disclosure relates to a methodfor treating a colon cancer in a mammal. This method involvesadministering, to the mammal, a therapeutically effective dose of ZnOparticles having substantially spherical shapes with diameters of 220nm-3.5 μm and BET surface areas of 8-25 m²/g.

In one embodiment, the therapeutically effective dose is 0.1 to 5 g ofZnO particles per kg of the mammal per day.

In one embodiment, the diameters are 1.5-3.5 μm, and the BET surfaceareas are 15-25 m²/g.

In one embodiment, the ZnO particles are porous with pore sizes of 20-35nm.

In one embodiment, a surface of the ZnO particles has nanopetals of50-200 nm thickness extending 100-500 nm from the surface, thenanopetals traversing the surface with lengths of 500 nm-2 μm.

In one embodiment, the ZnO particles are administered as a part of acomposition, wherein the composition further comprises a food product, apharmaceutically acceptable excipient, a pharmaceutically acceptablecarrier, or an antioxidant.

In one embodiment, the mammal is a human, a non-human primate, a dog, acat, a horse, a cow, a goat, a rat, a pig, a rabbit, or a mouse.

In one embodiment, the ZnO particles consist essentially of ZnO.

In one embodiment, the ZnO particles contact a first population of coloncancer cells in the colon cancer. At a time 24 hours after thecontacting, the first population has a growth inhibition of 60-80% inrelation to a second population of colon cancer cells in the coloncancer that were not contacted.

According to a second aspect, the present disclosure relates to a methodof reducing an organic contaminant concentration in an aqueous solution.This method involves contacting ZnO nanoflowers with the aqueoussolution comprising the organic contaminant at a contaminantconcentration of 1 mg/L-1 g/L and irradiating the ZnO nanoflowers whilein contact with the solution. Here, the ZnO nanoflowers have a generallyspherical shape with a diameter of 1.5-3.5 μm. A surface of the ZnOnanoflowers has nanopetals of 50-200 nm thickness extending 100-500 nmfrom the surface, and these nanopetals traverse the surface with lengthsof 500 nm -2 μm. The ZnO nanoflowers reduce the contaminantconcentration in the solution by adsorption and/or photocatalyticdegradation.

In one embodiment, the ZnO nanoflowers are dispersed within the solutionat a concentration of 0.5-100 mg/L.

In one embodiment, the irradiating is 15-30 minutes of UV lightirradiation, and the concentration of the organic contaminant after theirradiating is 40-50% of the concentration of the organic contaminantbefore the irradiating.

In one embodiment, the organic contaminant is at least one selected fromthe group consisting of pharmaceutical compound, a dye, a metabolite, amicrobial toxin, an herbicide, a pesticide, and a steroid.

In one embodiment, the ZnO nanoflowers are made by heating an aqueousZn²⁺ solution with sodium hydroxide.

In one embodiment, the aqueous Zn²⁺ solution comprises Zn(NO₃)₂.

In one embodiment, the ZnO nanoflowers consist essentially of ZnO.

In one embodiment, the ZnO nanoflowers have a band gap energy of2.90-3.31 eV.

In one embodiment, the irradiating uses sunlight as an irradiationsource.

In one embodiment, the method further comprises contacting the solutionwith a second absorbent or a second photocatalyst to reduce theconcentration of the contaminant and/or reduce a concentration of asecond contaminant in the solution.

In one embodiment, the method also involves rinsing the ZnO nanoflowersto produce cleaned ZnO nanoflowers and reusing the cleaned ZnOnanoflowers, which maintain a contaminant reduction capacity for atleast 5 purification cycles.

The foregoing paragraphs have been provided by way of generalintroduction, and are not intended to limit the scope of the followingclaims. The described embodiments, together with further advantages,will be best understood by reference to the following detaileddescription taken in conjunction with the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

A more complete appreciation of the disclosure and many of the attendantadvantages thereof will be readily obtained as the same becomes betterunderstood by reference to the following detailed description whenconsidered in connection with the accompanying drawings, wherein:

FIG. 1 is an X-ray diffraction pattern of ZnO nanoflowers and ZnOnanospheres.

FIG. 2A is an SEM image of ZnO nanoflowers, scale bar 10 μm.

FIG. 2B is a zoomed-in view of FIG. 2A, scale bar 5 μm.

FIG. 3A is an SEM image of ZnO nanoflowers, scale bar 10 μm.

FIG. 3B is a zoomed-in view of FIG. 3A, scale bar 5 μm.

FIG. 4 shows a UV-Vis diffuse reflectance spectrum of ZnO nanoflowersand ZnO nanospheres.

FIG. 5 shows N₂ adsorption-desorption isotherms of ZnO nanoflowers andZnO nanospheres.

FIG. 6 shows the photocatalytic activities of different amounts of ZnOnanoflowers over 120 minutes.

FIG. 7 shows the photocatalytic activities for reusing a 50 mg sample ofZnO nanoflowers.

FIG. 8 shows the photocatalytic activities of different amounts of ZnOnanospheres over 120 minutes.

FIG. 9 shows the photocatalytic activities for reusing a 50 mg sample ofZnO nanospheres.

FIG. 10 shows the changes in UV-Vis absorption spectra of methyl orangein the presence of 50 mg ZnO nanoflowers and with different irradiationtimes.

FIG. 11 shows the changes in UV-Vis absorption spectra of methyl orangein the presence of 50 mg ZnO nanospheres and with different irradiationtimes.

FIG. 12 shows the percent viability of HCT116 cells exposed to differentconcentrations of ZnO nanoflowers or ZnO nanospheres.

DETAILED DESCRIPTION OF THE EMBODIMENTS

Embodiments of the present disclosure will now be described more fullyhereinafter with reference to the accompanying drawings, in which some,but not all embodiments of the disclosure are shown.

The present disclosure will be better understood with reference to thefollowing definitions. As used herein, the words “a” and “an” and thelike carry the meaning of “one or more.” Within the description of thisdisclosure, where a numerical limit or range is stated, the endpointsare included unless stated otherwise. Also, all values and subrangeswithin a numerical limit or range are specifically included as ifexplicitly written out.

For polygonal shapes, the term “diameter,” as used herein, and unlessotherwise specified, refers to the greatest possible distance measuredfrom a vertex of a polygon through a central region of the face to thevertex on the opposite side. For a circle, an oval, and an ellipse,“diameter” refers to the greatest possible distance measured from onepoint on the shape through the centroid of the shape to a point directlyacross from it.

As used herein, “compound” is intended to refer to a chemical entity,whether as a solid, liquid, or gas, and whether in a crude mixture orisolated and purified.

In addition, the present disclosure is intended to include all isotopesof atoms occurring in the present compounds and complexes. Isotopesinclude those atoms having the same atomic number but different massnumbers. By way of general example and without limitation, isotopes ofhydrogen include deuterium and tritium. Isotopes of carbon include ¹³Cand ¹⁴C. Isotopes of oxygen include ¹⁶O, ¹⁷O, ¹⁸O, and others. Isotopesof zinc include, but are not limited to, ⁶⁴Zn, ⁶⁶Zn, ⁶⁷Zn, ⁶⁸Zn, and⁷⁰Zn. Isotopically-labeled compounds of the disclosure can generally beprepared by conventional techniques known to those skilled in the art orby processes analogous to those described herein, using an appropriateisotopically-labeled reagent in place of the non-labeled reagentotherwise employed. In addition, where compounds have hydration states,any degree or mixture of hydration states may be used.

The present disclosure relates to methods of using ZnO particles. Here,the ZnO particles have substantially spherical shapes with diameters of220 nm-3.5 μm, preferably 230 nm-500 nm, more preferably 250 nm-320 nm,or preferably 1.5 μm-3.5 μm, more preferably 2.2 μm-3.5 μm. As definedhere, the term “substantially spherical” means that the standarddeviation of the distance from anywhere on the outer surface to theparticle centroid (center of mass) varies by less than 30%, preferablyby less than 20%, more preferably by less than 10% of the averagedistance.

In one embodiment, the ZnO particles are monodisperse, having acoefficient of variation or relative standard deviation, expressed as apercentage and defined as the ratio of the particle diameter standarddeviation (σ) to the particle diameter mean (μ), multiplied by 100%, ofless than 25%, preferably less than 10%, preferably less than 8%,preferably less than 6%, preferably less than 5%. in a preferredembodiment, the ZnO particles are monodisperse having a particlediameter distribution ranging from 80% of the average particle diameterto 120% of the average particle diameter, preferably 85-115%, preferably90-110% of the average particle diameter. In another embodiment, the ZnOparticles are not monodisperse.

The ZnO particles may have BET surface areas of 8-25 m²/g, preferably15-24 m²/g, more preferably 18-23 m²/g, or preferably 8-15 m²/g, morepreferably 9-12 m²/g. Here, the surface area may be determined byBrunauer-Emmett-Teller (BET) analysis of N₂ adsorption isotherms, thoughother techniques may be used, such as mercury intrusion porosimetry.

In one embodiment, the ZnO particles are porous with pore sizes of 20-35nm, preferably 21-26 nm, more preferably 22-24 nm, or preferably 26-33nm, more preferably 28-32 nm. The ZnO particles may have pore volumes of0.05-0.16 cm³/g, preferably 0.06-0.11 cm³/g, more preferably 0.07-0.09cm³/g, or 0.10-0.15 cm³/g, more preferably 0.11-0.14 cm³/g.

In one embodiment, the ZnO particles may be present as agglomerates. Asused herein, the term “agglomerates” refers to a clustered particulatecomposition comprising primary particles, the primary particles beingaggregated together in such a way so as to form clusters thereof, atleast 50 volume percent of the clusters having a mean diameter that isat least 2 times the mean diameter of the primary particles, andpreferably at least 90 volume percent of the clusters having a meandiameter that is at least 5 times the mean diameter of the primaryparticles. The primary particles may be the ZnO particles having a meandiameter as those diameters previously described.

In one embodiment, the ZnO particles consist essentially of ZnO. Asdefined here, the ZnO particles consisting essentially of ZnO means that95-100%, preferably 96.0-99.7%, more preferably 97.5-99.5% of the massof ZnO particles is ZnO. Where the ZnO particles consist of less than100% ZnO, the ZnO particles may have adsorbed, reacted, or incorporatedcontaminants, for instance from gas molecules, other metals or metaloxides, or organic compounds. In an alternative embodiment, the ZnOparticles may be intentionally modified or mixed with other compounds.For example, the particles may be formed as a mixture of ZnS and ZnO ata molar ratio of 100:1-1:100, preferably 10:1-1:10, more preferably4:1-1:4. As another example, the surface of ZnO particles may bedecorated with other nanoparticles, for instance, CeO₂ or V₂O₅nanoparticles having diameters of 30-100 nm, preferably 40-90 nm, morepreferably 50-80 nm. In another embodiment, the ZnO particles maycomprise Zn metal.

The ZnO particles may comprise ZnO in the form an amorphous phase, acrystalline phase, or both. Crystalline ZnO may be in a hexagonalwurtzite phase or a cubic zincblende phase, where in both cases the zincand oxygen centers are tetrahedral. Preferably the ZnO is present in thewurtzite phase.

The ZnO particles may be further classified as ZnO nanospheres and ZnOnanoflowers based on size and morphology. The ZnO nanospheres havediameters of 220-500 nm, preferably 230-400 nm, more preferably 240-350nm and BET surface areas of 8-15 m²/g, preferably 8.5-12 m²/g, morepreferably 8.5-11 m²/g. The ZnO nanospheres may have pore sizes of 26-35nm, preferably 28-33 nm, more preferably 29-32 nm and pore volumes of0.05-0.11 cm³/g, preferably 0.06-0.10 cm³/g, more preferably 0.07-0.09cm³/g. The SEM images of FIGS. 3A and 3B show examples of ZnO particlesthat are ZnO nanospheres. In some embodiments, the ZnO nanospheres asdescribed above may be called “microspheres.”

The ZnO nanoflowers may have diameters of 1.5-3.5 μm, preferably 2.2μm-3.4 μm, even more preferably 2.3-3.3 μm, and BET surface areas of15-25 m²/g, preferably 18-24 m²/g, more preferably 20-23 m²/g. In otherembodiments, the ZnO nanoflowers may have smaller diameters, such asdiameters of 500 nm-1.5 μm, preferably 600 nm-1 μm, more preferably650-900 nm. The ZnO nanoflowers may have pore sizes of 20-26 nm,preferably 21-25 nm, more preferably 22-24 nm and pore volumes of0.10-0.15 cm³/g, preferably 0.11-0.14 cm³/g, more preferably 0.12-0.14cm³/g.

In one embodiment, a surface of the ZnO nanoflowers has nanopetals of50-200 nm thickness, preferably 70-150 nm thickness, more preferably85-120 nm thickness. In some embodiments, the nanopetals may instead becalled nanosheets or nanoplatelets. The nanopetals may extend 100-500 nmfrom the surface, preferably 150-400 nm from the surface, morepreferably 180-350 nm from the surface. The nanopetals may traverse thesurface with lengths of 500 nm-2 μm, preferably 800 nm-1.5 μm, morepreferably 900 nm-1.2 μm. In one embodiment, 70-100%, more preferably80-99%, more preferably 85-97% of the outer surface of the nanoflowersis covered with nanopetals. In alternative embodiments, a ZnO nanoflowermay have a lower coverage of nanopetals, or nanopetals that are shorteror longer in one or more dimensions. The SEM images of FIGS. 2A and 2Bshow examples of ZnO particles that are ZnO nanoflowers. In analternative embodiment, a ZnO particle that has a smaller diameter thana ZnO nanoflower may have nanopetals.

In some embodiments, the ZnO nanoflowers as described above may becalled “microflowers.”

In one embodiment, the ZnO nanoflowers and ZnO nanospheres have a bandgap energy of 2.90-3.31 eV, preferably 2.95-3.15 eV, more preferably3.00-3.12 eV, even more preferably 3.05-3.11 eV, though in someembodiments, the ZnO nanoflowers or ZnO nanospheres may have a band gapenergy less than 2.90 eV or greater than 3.31 eV. In one embodiment, theband gap energy is 3.07-3.09 eV, or about 3.08 eV. The band gap energymay be determined by UV-Vis adsorption or some other method.

In one embodiment, the ZnO particles, including both ZnO nanospheres andZnO nanoflowers, may be synthesized by heating an aqueous Zn²⁺ solutionwith sodium hydroxide. The sodium hydroxide (NaOH) may be present in theaqueous Zn²⁺ solution at a mass percentage of 0.5-1.5 mass %, preferably0.6-1.2 mass %, more preferably 0.9-1.1 mass %, relative to the combinedmass of aqueous Zn²⁺ solution and sodium hydroxide. The Zn²⁺ may bepresent at a mass percentage of 0.11-0.33 mass %, preferably 0.13-0.26mass %, more preferably 0.20-0.24 mass %, relative to the combined massof aqueous Zn²⁺ solution and sodium hydroxide. In alternativeembodiments, a different inorganic base may be used instead of NaOH, forinstance, Ca(OH)₂, KOH, or LiOH. The aqueous Zn²⁺ solution may be madeby mixing a zinc salt in water, for instance, ZnCl₂, Zn(NO₃)₂, ZnSO₄,ZnBr₂, or some other zinc salt. Preferably, the zinc salt is Zn(NO₃)₂.In one embodiment, the Zn²⁺ comes from mixing Zn(NO₃)₂ in water,including Zn(NO₃)₂ originally in an anhydrous form or a hydrated form(e.g. Zn(NO₃)₂.6H₂O).

To synthesize ZnO nanospheres having the sizes and morphologies asdescribed previously, the mixture of aqueous Zn²⁺ solution and. NaOH maybe heated in an autoclave for 6-48 hours, preferably 12-36 hours, morepreferably about 24 hours, at a temperature of 100-180° C., preferably120-160° C., more preferably 130-150° C. to form precipitated ZnO. Thereaction solution may be stirred during the heating. Preferably theautoclave surface in contact with the solution is coated with orcomprises a non-reactive material, such as PTFE. The solution may becooled to room temperature or about 20-28° C., preferably 23-27° C. TheZnO may be recovered from the solution by filtration and/orcentrifugation, and washed with water and/or an alcohol. The washed ZnOmay be dried in an oven for 16-32 hours, preferably 20-28 hours at atemperature of 50-80° C., preferably 55-70° C., to form ZnO nanospheres.

To synthesize ZnO nanoflowers having the sizes and morphologies asdescribed previously, the mixture of aqueous Zn²⁺ solution and NaOH maybe heated or refluxed for 3-7 hours, preferably 4-6 hours, morepreferably about 5 hours, at a temperature of 60-100° C., preferably70-90° C., more preferably 75-85° C. to form precipitated ZnO.Preferably the reaction solution is stirred during the heating orrefluxing. The solution may be cooled to room temperature or about20-28° C., preferably 23-27° C. The ZnO may be recovered from thesolution by filtration and/or centrifugation, and washed with waterand/or an alcohol. The washed ZnO may be dried in an oven for 16-32hours, preferably 20-28 hours at a temperature of 50-80° C., preferably55-70° C. to form ZnO nanoflowers.

In alternative embodiments, ZnO nanospheres or ZnO nanoflowers havingsizes and morphologies as described above may be synthesized throughcompletely different processes.

According to a first aspect, the present disclosure relates to a methodfor treating a colon cancer in a mammal. This method involvesadministering a therapeutically effective dose of ZnO particles. The ZnOparticles may have the sizes and morphologies as described above, andmay be present as ZnO nanoflowers and/or ZnO nanospheres.

In a preferred embodiment, the cancer is colon cancer. However, in otherembodiments, the cancer may be breast cancer, liver cancer, colorectalcancer, stomach cancer, skin cancer, prostate cancer, ovarian cancer,testicular cancer, renal cancer, brain cancer, lung cancer, uterinecancer, bladder cancer, esophageal cancer, or pancreatic cancer. Thecancer may be an adenocarcinoma, a basal cell carcinoma, a squamous cellcarcinoma, a renal cell carcinoma, a ductal carcinoma in situ (DCIS), aninvasive ductal carcinoma, a transitional cell carcinoma, a soft tissuesarcoma, or leukemia. In one embodiment, the mammal is a human, anon-human primate, a dog, a cat, a horse, a cow, a goat, a rat, a pig, arabbit, or a mouse. Preferably the mammal is a human.

A “therapeutically effective dose” refers to an amount of the ZnOparticles being administered which will relieve to some extent one ormore of the symptoms of the cancer being treated. In another embodiment,a “therapeutically effective dose” refers to the amount which has theeffect of inhibiting (that is, slowing to some extent, or preferablystopping) cancer cell growth or proliferation. Similarly, the phrase“treat a cancer in a mammal” refers to administering a therapeuticallyeffective dose of the ZnO particles to a mammal.

As used herein, the terms “treat”, “treatment”, and “treating,” in thecontext of the administration of a therapeutically effective dose of theZnO particles to a mammal, refer to the reduction or inhibition of theprogression and/or duration of a cancer, the reduction or ameliorationof the severity of the cancer, and/or the amelioration of one or moresymptoms thereof resulting from the administration of one or moretherapies. “Treating” or “treatment” of the cancer includes preventingthe cancer from occurring in a subject that may be predisposed to thecancer but does not yet experience or exhibit symptoms of the cancer(prophylactic treatment), inhibiting the cancer (slowing or arrestingits development), ameliorating the cancer, providing relief from thesymptoms or side-effects of the cancer (including palliative treatment),and relieving the cancer (causing regression of the cancer). With regardto the cancer, these terms simply mean that one or more of the symptomsof the cancer will be reduced. Such terms may refer to one, two, three,or more results following the administration of one, two, three, or moretherapies: (1) a stabilization, reduction (e.g. by more than 10%, 20%,30%, 40%, 50%, preferably by more than 60% of the population of cancercells and/or tumor size before administration), or elimination of thecancer cells, (2) inhibiting cancerous cell division and/or cancerouscell proliferation, (3) relieving to some extent (or, preferably,eliminating) one or more symptoms associated with a pathology related toor caused in part by unregulated or aberrant cellular division, (4) anincrease in cancer-free, relapse-free, progression-free, and/or overallsurvival, duration, or rate, (5) a decrease in hospitalization rate, (6)a decrease in hospitalization length, (7) eradication, removal, orcontrol of primary, regional and/or metastatic cancer, (8) astabilization or reduction (e.g. by at least 10%, 20%, 30%, 40%, 50%,60%, 70%, preferably at least 80% relative to the initial growth rate)in the growth of a tumor or neoplasm, (9) an impairment in the formationof a tumor, (10) a reduction in mortality, (11) an increase in theresponse rate, the durability of response, or number of patients whorespond or are in remission, (12) the size of the tumor is maintainedand does not increase or increases by less than 10%, preferably lessthan 5%, preferably less than 4%, preferably less than 2%, (13) adecrease in the need for surgery (e.g. colectomy, mastectomy), and (14)preventing or reducing (e.g. by more than 10%, more than 30%, preferablyby more than 60% of the population of metastasized cancer cells beforeadministration) the metastasis of cancer cells.

In one embodiment, the ZnO particles are administered as a part of acomposition, wherein the composition further comprises a food product, apharmaceutically acceptable excipient, a pharmaceutically acceptablecarrier, or an antioxidant.

In one embodiment of the method, the composition may comprise a foodproduct. The food product may be an ingredient to improve a flavor orappearance of the ZnO particles for oral administration, such as sugar,food coloring, non-nutritive sweeteners, preservatives, artificialflavoring, or natural flavoring. In one embodiment, the food product maybe a snack or candy, such as dried fruit, a lozenge, fruit leather,yogurt, pudding, a gummy, an energy bar, a candy bar, or a chewing gum.In other embodiments, the food product may be a drink, such as tea,water, milk, smoothie, soft drink, or shake. In other embodiments, thefood product may be a food that is part of a meal. In one embodiment,the food product may comprise 0.01-50 mass %, preferably 0.1-10 mass %,more preferably 0.2-1.0 mass % ZnO particles relative to a combined massof the ZnO particles and food product. However, in some embodiments, thefood product may comprise less than 0.01 mass % or more than 50 mass %ZnO particles.

In one embodiment, the composition comprises 1-99.9%, preferably10-99.9%, more preferably 20-99.9%, more preferably 30-99.9%, morepreferably 40-99.9%, more preferably 50-99.9%, more preferably 60-99.9%,more preferably 70-99.9%, more preferably 80-99.9%, even more preferably90-99.9% of ZnO particles, and 0.1% or more of the pharmaceuticallyacceptable carrier or excipient, based on the total weight of thecomposition.

In one embodiment, the pharmaceutically acceptable carrier or excipientis a dispersing agent, a disintegrating agent, a binding agent, or alubricating agent.

In one embodiment, the pharmaceutically acceptable carrier or excipientmay be a sugar, such as lactose, glucose, or sucrose; cellulose, or itsderivatives, such as sodium carboxymethyl cellulose, ethyl cellulose andcellulose acetate; starches, such as corn starch, tapioca starch, andpotato starch; crospovidone, croscarmellose sodium, sodium starchglycolate, gelatin, alginate, carnauba wax, pregelatinized starch,xylitol, mannitol, sorbitol, polyethylene glycol, polyvinylpyrrolidone,talc, silica, sodium stearyl fumarate, magnesium stearate, stearic acid,sodium benzoate, sodium lauryl sulfate, mineral oil, palmitic acid, ormixtures thereof.

In one embodiment, the composition is formulated for local or systemiceffect, and may be administered by topical, enteral, or parenteralroutes. Modes of administration may include, but are not limited to,transdermal administration, eye drops, ear drops, oral administration,intravenous administration, topical administration, inhalation spray,rectal administration, intradermal administration, transdermaladministration, subcutaneous administration, intramuscularadministration, intralesional administration, intrapulmonaladministration, epidural administration, intravesical administration,intracranial administration, intracardial administration, intrasternaladministration and sublingual administration.

In one embodiment the composition is in solid, semi-solid, or liquiddosage forms. In solid dosage forms for oral administration (includingbut not limited to capsules, tablets, pills, powders, and granules), thecomposition is mixed with one or more pharmaceutically acceptablecarriers or excipients such as: (1) fillers or extenders, such asstarches, lactose, sucrose, glucose, mannitol, and silicic acid; (2)binders, such as carboxymethylcellulose, alginate, gelatin, polyvinylpyrrolidone, and sucrose; (3) humectants, such as glycerol; (4)disintegrating agents, such as alginate, calcium carbonate, potato ortapioca starch, silica, and sodium carbonate; (5) solution retardingagents, such as paraffin; (6) absorption accelerators, such asquaternary ammonium compounds; (7) wetting agents, such as acetylalcohol and glycerol monostearate; (8) absorbents, such as kaolin andbentonite clay; (9) lubricants, such a talc, calcium stearate, magnesiumstearate, solid polyethylene glycols, sodium lauryl sulfate, andmixtures thereof; and (10) coloring agents. In the case of capsules,tablets and pills, the compositions may also comprise buffering agents.

Semi-solid dosage forms may be used for topical administration and mayinclude sprays, ointments, pastes, creams, lotions, gels, and patches.These forms may further include pharmaceutically acceptable carriers orexcipients such as animal and vegetable fats, oils, waxes, paraffins,starch, tragacanth, cellulose derivatives, polyethylene glycols,silicones, bentonites, silicic acid, talc, and zinc oxide.

Liquid dosage forms for oral administration include pharmaceuticallyacceptable emulsions, microemulsions, solutions, suspensions, syrups,and elixirs. The liquid dosage forms may also contain inert diluentscommonly used in the art, such as, for example, water or other solvents,solubilizing agents and emulsifiers, such as ethyl alcohol, isopropylalcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzylbenzoate, propylene glycol, 1,3-butylene glycol, oils (in particular,cottonseed, corn, peanut, sunflower, soybean, olive, castor, and sesameoils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fattyacid esters of sorbitan, and mixtures thereof.

In one embodiment of the method, the ZnO particles are administered as apart of a composition which further comprises an antioxidant. Theantioxidant may be melatonin, lutein, α-carotene, β-carotene,astaxanthin, tocotrienol, tocopherol, ascorbic acid, gallic acid,ellagic acid, or lycopene. The weight ratio of the antioxidant to theZnO particles may be 1:65,000-1:1, preferably 1:10,000-1:100, morepreferably 1:5,000-1:500. However, in other embodiments, the antioxidantmay be administered at a greater mass than the ZnO particles.

In one embodiment, the therapeutically effective dose is 0.1-5 g,preferably 1.0-4.0 g, more preferably 1.5-2.5 g of ZnO particles per kgof the mammal per day. In another embodiment, the therapeuticallyeffective dose may be 0.1-1 g, 1-2 g, 2-3 g, 3-4 g, or 4-5 g of ZnOparticles per kg of the mammal per day. In other embodiments, thetherapeutically effective dose may be less than 0.1 g or more than 5 gof ZnO particles per kg of the mammal per day. In one embodiment, theadministration of a therapeutically effective dose of ZnO particles maybe split or distributed over a single day. For example, a 1 g per kg perday dose could be split into two separate doses of 0.5 g per kg, andadministered at two different times in a day, for example, 8 AM and 7PM. Or, a 1 g per kg per day dose could be split into four separatedoses of 0.25 g per kg, and administered at four different times in aday. In a related embodiment, a daily dose may be a combined doseadministered less frequently than every day. For example, a 0.75 g perkg per day dose could be administered as a 1.5 g per kg dose every otherday. In certain embodiments, the dose may be continually administeredthroughout an entire day or a part of a day, such as by intravenousadministration. Where a dose is administered continuously throughoutpart of the day, it may be administered for 2-18 h, or 4-16 h.

A person having ordinary skill in the art may be able to adjustadministration based on the changes in expression levels of one or morebiomolecules. These biomolecules may be measured from a patient's bloodserum. These biomolecules include, but are not limited tocarcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (CA19-9),MAPKAPK3, ACVR2B, and/or circulating DNA markers for K-ras mutations.For example, a human patient with colon cancer who is administered theZnO particles at a dosage of 1 g per kg bodyweight per day for 3 monthsmay show serum levels of CEA decrease by 15% as compared to before theadministering. The daily dosage may be increased to 2 g per kgbodyweight to cause a desired 30-70% decrease in CEA serum levels. Anormal serum concentration of CEA is ≤3 ng/mL, and for example, a cancerpatient may have a concentration of about 10 or about 30 ng/mL. In otherembodiments, a percent change in enzyme activity or biomoleculeexpression level may warrant modifications to other treatment methods,such as other supplements, chemotherapy, or other cancer treatments.

In one embodiment, the ZnO particles may be administered in conjunctionwith other forms of cancer treatment, such as radiation therapy orchemotherapy. Preferably the ZnO particles may be administered 1-4 weeksor 2-3 weeks before starting a chemotherapy regimen, and theadministration would continue throughout the duration of thechemotherapy. The administering may be stopped at the same time as theother cancer treatment, or the ZnO particles may be administered for alonger period. It is envisioned that chemotherapy drugs may worksynergistically with the ZnO particles. In one embodiment, the ZnOparticles may be used as drug delivery vehicles, for example, to deliverand release compounds of a FOLFIRI-Bevacizumab treatment (chemotherapyregimen comprising ceucovorin calcium (folinic acid), fluorouracil,irinotecan hydrochloride, and bevacizumab (AVASTIN)).

In an alternative embodiment, cancer cells may be treated with the ZnOparticles in vitro, for instance, as a way to test the ZnO particlesunder different conditions in a controlled environment or withadditional drugs. These cancer cells may come from a biopsy of a mammal,for instance a biopsy of a colon cancer, or the cells may be from anestablished cancer cell line, for instance, HCT 116, MDA-MB-231, MCF-7,AU565, BT20, HeLa, HepG2, SNU-475, LH86, Caco-2, NCI-H250, A-498, Eph41424.2, SK-MES-1, DU 145, CHLA-02-ATRT, SCC-4, A-253, SNU-C2B, LS513, orsome other cancer cell line. The cells may come from a cancer thatformed on its own in a mammal, or may come from a cancer that was formedby chemical induction or radiation. Diethyl nitrosoamine, (DEN),7,12-dimethylbenz[a]anthracene (DMBA),12-O-tetradecanoylphorbol-13-acetate (TPA), azoxymethane (AOM), or someother carcinogenic compound may be used to chemically induce cancer.Additionally, the cancer cells may be derived from a tumor or cancercells that were transplanted and allowed to grow in a mammal.

In one embodiment of the method, the ZnO particles contact a firstpopulation of colon cancer cells in the colon cancer. At a time 24 hoursafter the contacting, the first population has a growth inhibition of60-80%, preferably 65-78%, more preferably 70-77% in relation to asecond population of colon cancer cells in the colon cancer that werenot contacted. However, in other embodiments, the growth inhibition maybe lower than 60% or greater than 80%. As defined here, the growthinhibition is the percentage decrease in viability of the cellpopulation upon treatment. The viability of a cell population is thepercentage of living cells in relation to a second control cellpopulation. The viability may be measured by a standard live/dead cellassay, such as an MTT assay, flow cytometry, by dyeing and manuallycounting cells, or by some other method. The viability may also bedetermined at a time earlier or later than 24 hours after thecontacting.

In an alternative embodiment, given the photocatalytic activity of theZnO particles, the method may further involve a step of irradiating theZnO particles in contact with the colon cancer cells. This may be a typeof photodynamic therapy (PDT).

According to a second aspect, the present disclosure relates to a methodof reducing a concentration of an organic contaminant in an aqueoussolution. This method involves contacting ZnO nanoflowers with theaqueous solution comprising the contaminant and irradiating the ZnOnanoflowers while in contact with the aqueous solution. The ZnOnanoflowers reduce the contaminant concentration in the aqueous solutionby adsorption and/or photocatalytic degradation.

In one embodiment, the organic contaminant is at least one selected fromthe group consisting of pharmaceutical compound, a dye, a metabolite, amicrobial toxin, an herbicide, a pesticide, and a steroid. In apreferred embodiment, the organic contaminant may be a dye. In a furtherembodiment, where the organic contaminant is a dye, the dye may be anazin dye, an azo dye, a diarylmethane dye, a fluorescent dye, a foodcoloring, a fuel dye, an ikat dye, an indigo structured dye, anindophenol dye, a perylene dye, a phenol dye, a quinoline dye, arhodamine dye, a solvent dye, a staining dye, a thiazine dye, a thiazoledye, a triarylmethane dye, a vat dye, a violanthrone dye, or some othertype of dye. In a preferred embodiment,the dye may be a azo dye, inparticular, methyl orange (sodium4-{[4-(dimethylamino)phenyl]diazenyl}benzene-1-sulfonate).

The organic contaminant may be present in the aqueous solution at aconcentration of 1 mg/L-1 g/L, preferably 2 mg/L-500 mg/L, morepreferably 5 mg/L-200 mg/L, though in some embodiments, the organiccontaminant may be present in the aqueous solution at a concentration ofless than 1 mg/L or greater than 1 g/L.

The aqueous solution may come from an ocean, a bay, a river, a lake, aswamp, a pond, a pool, a fountain, a bath, an aquarium, a watertreatment plant, a sewage treatment plant, a desalination plant, amanufacturing plant, a chemical plant, a textile plant, a power plant, agas station, a food processing plant, a restaurant, a dry cleaner, orsome other place that may generate contaminated water mixtures. Inanother embodiment, the aqueous solution may be prepared in a laboratoryor pilot plant for the purpose of testing contaminant removal. In someembodiments, the aqueous solution may be a brine, or comprise sea wateror salt water.

In one embodiment, the aqueous solution may comprise a non-polar liquidphase at a volume percent concentration of 0.5-50%, preferably 2-40%,more preferably 4-30% relative to a total volume of the contaminatedwater mixture. The non-polar liquid phase may be emulsified or dispersedthroughout the aqueous solution, may float at the top of the aqueoussolution, or some combination of both. In another embodiment, theaqueous solution may not contain a non-polar liquid phase.

In one embodiment, the ZnO nanoflowers may be contacted with the aqueoussolution for a period of time effective to reduce the contaminantconcentration by 40-100%, preferably 50-99%, more preferably 60-95%. Inone embodiment, this time may be 20-180 minutes, preferably 30-120minutes, more preferably 40-105 minutes, though in some embodiments thetime may be shorter than 20 minutes or longer than 180 minutes. In oneembodiment, the ZnO nanoflowers may be contacted with the aqueoussolution by dispersing the ZnO nanoflowers in a fixed volume of aqueoussolution, and then stirring or agitating the aqueous solution to keepthe ZnO nanoflowers evenly mixed throughout. In one embodiment, the ZnOnanoflowers are dispersed within the aqueous solution at a concentrationof 0.5-100 mg/L, preferably 5-80 mg/L, more preferably 10-60 mg/L,though in some embodiments, the ZnO nanoflowers may be dispersed withinthe aqueous solution at a concentration of less than 0.5 mg/L or greaterthan 100 mg/L.

In one embodiment, the ZnO nanoflowers may not be dispersed in aqueoussolution but fixed to a solid support, such as a plate or a wire mesh.In one embodiment, the solid support may be planar so that irradiatedlight may more evenly spread on the ZnO nanoflowers. The solid supportmay also be a single piece so that the ZnO nanoflowers can be easilyremoved from the aqueous solution, or removed from a vessel. In afurther embodiment, where the ZnO nanoflowers are attached to a solidsupport so that they do not disperse, the aqueous solution may becontinually flowed over the ZnO nanoflowers while irradiating. Inanother related embodiment, the aqueous solution may be intermittentlyflowed over the ZnO nanoflowers while irradiating. Alternatively, theZnO nanoflowers may be dispersed but confined within a volume of wiremesh. In another embodiment, the ZnO nanoflowers may be fixed to a solidsupport, but dispersed in aqueous solution. For example, the ZnOnanoflowers may be attached to magnetic microparticles having diametersof 10-400 μm, preferably 40-200 μm.

In one embodiment, the ZnO nanospheres may be used instead of ZnOnanoflowers in the second aspect of the disclosure for reducing thecontaminant concentration in the aqueous solution by adsorption and/orphotocatalytic degradation.

In an alternative embodiment, ZnO particles larger than ZnO nanoflowers,or smaller than the ZnO nanospheres, may be used in a similar procedureor arrangement for reducing the contaminant concentration in the aqueoussolution by adsorption and/or photocatalytic degradation.

Preferably, the illuminating is with UV light. The UV light source maybe a mercury or xenon gas discharge lamp, an electric arc, sunlight, alight emitting diode (LED), a laser, a fluorescent lamp, a cathode raytube, or some other source. In one embodiment, filters, reflectors,collimators, fiber optics, polarizers, and/or lenses may be used tomanipulate the light path or properties of the light from the lightsource. For example, one or more reflectors may be used to focus thelight from a mercury gas discharge lamp onto ZnO nanoflowers fixed on asubstrate or into an aqueous solution having dispersed ZnO nanoflowers.Alternatively, a reflector may be positioned opposite the light sourcein order to reflect stray UV light back towards the aqueous solution. Inone embodiment, two or more light sources may be used, which may be ofthe same type or different types, and may be positioned on the same sideor on different sides of the aqueous solution. As another example, wheresunlight is used as a light source, the sunlight may be filtered,reflected, and focused into the aqueous solution to increase theproportion of UV light intensity while minimizing heating and radiationfrom other wavelengths. For instance, a Wood's glass optical filter maybe used to allow UV light to pass while blocking other wavelengths. Inone embodiment, a UV light source may also emit wavelengths longer thanthe UV range of 100-400 nm range, for instance, wavelengths of 405-420nm.

In one embodiment, the UV light has an intensity of 450-1550 mW/cm²,preferably 600-1400 mW/cm², more preferably 800-1200 mW/cm². However, inalternative embodiments, a lower intensity may be used if the flow rateis slowed or if the filtered water product is reapplied to the feed sidecoating. The UV light source may emit light within the wavelength rangeof 100-410 nm, preferably 370-405 nm, more preferably 390-400 nm.Depending on the composition and morphology of the photocatalyst used,certain UV wavelengths may be more preferable than others. Ideally, theUV wavelength corresponds to an energy equal to or greater than theelectronic band gap energy of the ZnO nanoflowers.

In one embodiment, the ZnO nanoflowers reduce the contaminantconcentration in the aqueous solution by photocatalytic degradation.Here, exposure of the ZnO nanoflowers to an irradiation of a wavelengthcorresponding to the band gap energy or a greater energy may cause thephotoexcitation of ZnO electrons into a conduction band with acorresponding generation of holes in a valence band. The strongreduction power of the electrons and the strong oxidation power of theholes may lead to the decomposition of organic materials, preferablyinto harmless byproducts, which produces a reduced concentration oforganic contaminant. In one embodiment, the ZnO nanoflowers may causeother reactions, such as hydrolysis and/or water splitting.

In one embodiment, a light source may be located outside of a vesselcontaining the aqueous solution, and may transmit UV light and/or otherwavelengths through an additional opening in the vessel wall or througha transparent window in the vessel wall. For example, the transparentwindow may comprise quartz or a polymeric material transparent to UVlight such as poly(methyl methacrylate) (also known as PLEXIGLAS). Asdefined herein, “transparent” refers to an optical quality of a compoundwherein a certain wavelength or range of wavelengths of light maytraverse through a portion of the compound with a small loss of lightintensity. Here, the “transparent window” may causes a loss of less than10%, preferably less than 5%, more preferably less than 2% of theintensity of a wavelength of UV light. In one embodiment, the vesselwall and transparent window may comprise the same material, for example,a vessel may comprise poly(methyl methacrylate) walls, which may alsofunction as transparent windows. In another embodiment, a vessel wallmay have a window that is partially transparent to UV light, forinstance, a window comprising soda lime glass.

Where an irradiation source emits heat, the aqueous solution or a vesselcontaining the aqueous solution may be temperature-regulated to preventoverheating and/or evaporation, for example, by water tubing, a waterand/or ice bath, ice packs, heat sinks, or by air cooling. Devices tomeasure and record the physical and/or chemical properties of theaqueous solution may be submerged in the aqueous solution or connectedthrough a wall of a vessel containing the aqueous solution. Examples ofthese devices include, but are not limited to, pressure gauges,flowmeters, conductivity meters, pH meters, temperature sensors, andspectrophotometers.

In one embodiment, the ZnO nanoflowers may reduce the organiccontaminant concentration by adsorption. Here, molecules of the organiccontaminant adhere to the surface of the ZnO nanoflowers, on an outerexposed surface or within the nanopetals or pores. The adsorption mayresult from electrostatic attraction, physisorption, and/orchemisorption. In one embodiment, the ZnO nanoflowers may reduce theorganic contaminant concentration only by adsorption. For instance, thiscondition may occur if the ZnO nanoflowers and aqueous solution are notexposed to light. In another embodiment, the ZnO nanoflowers may reducethe organic contaminant concentration substantially throughphotocatalytic degradation, for instance, under high intensity UV lightor with short contact times. In another embodiment, the ZnO nanoflowersmay reduce the organic contaminant concentration by both adsorption andphotocatalytic degradation. In some cases, contaminant molecules mayfirst adsorb to ZnO nanoflowers for a moment before beingphotocatalytically degraded. In other cases, a combination of bothadsorption and photocatalytic degradation may cause a reduction incontaminant concentration. For example, 30-90%, preferably 40-80% of thetotal moles of contaminant removed from the aqueous solution may be aresult of photocatalytic degradation, with the remaining moles beingremoved by adsorption. In one embodiment, sunlight may be used as theirradiation source in order to reduce the use of electricity.

The aqueous solution may or may not be pre-processed, for instance, byfiltering through a coarse filter to remove large particulate matter, orby exposure to UV light or ozone. In one embodiment, the method furthercomprises contacting the aqueous solution with a second absorbent or asecond photocatalyst to reduce the concentration of the contaminantand/or reduce a concentration of a second contaminant in the aqueoussolution. This contacting with a second absorbent or photocatalyst maybe done before, during, or after the step of mixing the ZnO nanoflowerswith the aqueous solution. In one embodiment, the ZnO nanoflowers andthe second absorbent or photocatalyst may be present in the same volumeof aqueous solution simultaneously. In another embodiment, one may bemixed and then removed before the second is added. In anotherembodiment, one may be mixed, and then the other may be mixed together.In another embodiment, both ZnO nanoflowers and second absorbent orphotocatalyst may be added and mixed together at the same time. Thesecond absorbent or photocatalyst may be dispersed in the aqueoussolution, fixed to a solid support, or confined on a packed bed. Inanother embodiment, an adsorbent may be used in place of the secondabsorbent.

In one embodiment, the irradiating is carried out for 15-30 minutes,preferably 18-28 minutes, more preferably 20-25 minutes, and thecontaminant concentration after the irradiating is 40-50%, preferably42-48%, or about 45% of the contaminant concentration before theirradiating. In a further embodiment, this irradiating is by UV lightirradiation. In other embodiments, the irradiating may be carried outfor shorter or longer times, and the contaminant concentration may beless than 40% or greater than 50% of the original concentration. Thesevariations may depend on parameters including but not limited to theintensity and wavelength of the irradiation, the type of contaminant,the concentration of contaminant and/or ZnO nanoflowers, and the surfacearea of the ZnO nanoflowers. In an alternative embodiment, the UV lightirradiation may directly photobleach, react, or degrade a contaminantwithout the contaminant having to contact the ZnO nanoflowers.

In one embodiment, the contaminant concentration may be determined bycomparing the change in the UV-Vis absorbance at certain wavelengths.For example and without limitation, a contaminant that absorbs stronglyat a wavelength of 465 nm may have its relative concentration changedetermined by observing the decrease in 465 nm absorbance. In a furtherembodiment, more than one wavelength may be monitored.

In one embodiment, the method also involves rinsing the ZnO nanoflowersto produce cleaned. ZnO nanoflowers and reusing the cleaned ZnOnanoflowers, which maintain a contaminant reduction capacity for atleast 5 purification cycles. In one embodiment, the ZnO nanoflowers maybe removed or separated from the aqueous solution prior to the rinsing,by filtering, centrifugation, evaporation, decanting, or some othermeans. In one embodiment, the ZnO nanoflowers may be attached to amagnetic solid support, such as Fe microparticles, and the ZnOnanoflowers may be removed from aqueous solution by a permanent magnetor an electromagnet. Where the ZnO nanoflowers may be attached to asingle solid support, such as a plate or a wire mesh, the entire supportmay be lifted from the aqueous solution, or the aqueous solution drainedaway from the support. Preferably the ZnO nanoflowers are rinsed withwater, such as deionized or distilled water, and in some embodiments,the ZnO nanoflowers may be rinsed with an organic solvent such aschloroform, acetone, methanol, or ethanol. In a preferred embodiment,the ZnO nanoflowers are rinsed with deionized water. In one embodiment,the ZnO nanoflowers may be sonicated in water or some other solvent.Where the ZnO nanoflowers are rinsed with an organic solvent, preferablythe ZnO nanoflowers are subsequently dried or rinsed with water in orderto remove the organic solvent.

As used herein, “maintains a contaminant reduction capacity” means thatunder substantially identical conditions (ZnO nanoflower concentration,contact time, contaminant type and concentration, irradiation, etc.)reused ZnO nanoflowers are able to decrease a contaminant concentrationby an amount that is within 70%, preferably within 80%, more preferablywithin 90% of its initial decrease in contaminant concentration. In someinstances, the ZnO nanoflowers may have greater contaminant reductioncapacities given longer contact times.

A purification cycle refers to the adsorption and/or photocatalyticdegradation of a contaminant by the ZnO nanoflowers and the subsequentrinsing of the ZnO nanoflowers to produce cleaned ZnO nanoflowers.Preferably the cleaned ZnO nanoflowers are able to maintain theircontaminant reduction capacity across different contaminants. In otherembodiments, the cleaned ZnO nanoflowers are able to maintain theiradsorption capacity for at least 5 cycles, at least 10 cycles, at least20 cycles, or even at least 50 cycles.

Preferably in reusing the ZnO nanoflowers, all of it may be recoveredafter each purification cycle, enabling several cycles to be repeatedwith a single batch of ZnO nanoflowers. However, in some embodiments,0.1-1 mass %, or 1-5 mass % may be lost with each cycle. Preferably thereuse of the ZnO nanoflowers does not change its morphology or otherphysical characteristics.

Where the ZnO nanoflowers are fixed to single support and exposed to aflowing aqueous solution comprising the aqueous organic contaminant, theused ZnO nanoflowers may be rinsed in place and optionally dried whilestaying fixed to the support. Alternatively, the ZnO nanoflowers may notbe fixed to a support, but confined within a permeable membrane orfilter, allowing similar operation.

In one embodiment, all of the above method and characteristics ofcontaminant reduction may be applied in full or in part to the ZnOnanospheres of the present disclosure.

The examples below are intended to further illustrate protocols forpreparing, characterizing, and using the zinc oxide nanospheres andnanoflowers, and are not intended to limit the scope of the claims.

EXAMPLE 1 Preparation of Zinc Oxide (ZnO) Nanoflowers

0.2 g of zinc nitrate (Zn(NO₃)₂.6H₂O) was weighed and transferred into around bottomed flask with 20 mL deionized water and stirred at roomtemperature. This was followed by the addition of 0.2 g sodium hydroxide(NaOH) to the flask, which was then stirred for 10 minutes at roomtemperature. The mixture was then refluxed at 80° C. for 5 hours. Theflask was cooled to room temperature and the precipitate wascentrifuged, washed with deionized water, and then washed with methanol.The product was dried in an oven for 24 hours at 60° C.

EXAMPLE 2 Preparation of Zinc Oxide (ZnO) Nanospheres

0.2 g of zinc nitrate (Zn(NO₃)₂.6H₂O) was weighed and transferred into aTEFLON-lined autoclave, followed by addition of water (20 mL) and 0.2 gof sodium hydroxide (NaOH). The mixture was stirred at room temperatureand then heated at 140° C. for 24 hours. The product was cooled to roomtemperature, and the precipitate was centrifuged, washed with deionizedwater, and then washed with methanol. The product was dried at 60° C. inan oven for 24 hours.

EXAMPLE 3 Physical Characterization

The morphologies of the zinc oxide nanoflowers and nanospheres wereexamined by scanning electron microscopy (SEM, FEI INSPECT S50), asshown in FIGS. 2A-2B and FIGS. 3A-3B, respectively. The crystallinityand crystal phases of the zinc oxide nanoflowers or nanospheres werestudied by an X-ray diffractometer (XRD Rigaku, Japan) using Cu-Kαradiation (λ=1.5418 {acute over (Å)}) in the range of 10°-80° with1°/min scanning speed. The XRD patterns for both nanoflowers andnanospheres arc shown in FIG. 1. UV-Vis diffuse reflectance spectra ofzinc oxide nanoflowers and nanospheres, as shown in FIG. 4, wererecorded on a diffuse reflectance UV-Vis spectrophotometer (JASCOV-750). Micromeritics ASAP 2020 PLUS nitrogen adsorption apparatus (USA)was employed for BET surface area determination. Before surface areaanalysis, samples were degassed at 180° C., and the surface area wasthen determined using N₂ adsorption data in the relative pressure (P/P₀)range of 0.05-0.3, as shown in FIG. 5. The BET surface area of zincoxide nanoflowers was observed as 22.54 m²/g (pore size: 22.92 nm; porevolume 0.1291 cm³/g) while for nanospheres, the surface area was 9.51m²/g (pore size: 30.74 nm; pore volume 0.0780 cm³/g).

EXAMPLE 4 Photocatalytic Activity

The photocatalytic activity of zinc oxide nanoflowers and nanosphereswas evaluated through the photocatalytic degradation of methyl orangeunder UV light irradiation for 120 min using a xenon lamp (300 W) as thelight source. In each experiment, 10 mg, 30 mg, and 50 mg of catalyst(zinc oxide nanoflowers or nanospheres) was dispersed in 100 mL of anaqueous solution of methyl orange dye, the dye having a concentration of10 mg/L. In order to ensure the adsorption-desorption equilibriumbetween catalyst and dye, the solution was stirred in the dark for 1 hbefore irradiating with the xenon lamp. At 15 min time intervals, 4 mLsamples of the suspension were collected and centrifuged to remove thezinc oxide nanoflowers or nanospheres, and the concentration of dye inthe supernatant was assessed with a UV-Visible spectrophotometer (JASCOV-750) by measuring the absorbance at 465 nm. The change in 200-800 nmabsorbance over time is shown in FIGS. 10 and 11. The degradationefficiency was calculated as:

Degradation efficiency (%)=(C ₀ −C)/C ₀×100%,

where C₀ is the initial concentration of the methyl orange dye, and C isthe time-dependent concentration of methyl orange upon irradiation. Thechange in degradation efficiency over time is shown in FIGS. 6-9.

EXAMPLE 5 Cell Culture and Cytotoxicity Activity

The human colon cancer cell line HCT116 was purchased from ATTC(American Type Culture Collection, USA) and maintained in DMEM medium.The cytotoxicities of zinc oxide nanoflowers and nanospheres wereevaluated against HCT116 cells by MTT assay. The MTT assay was performedwith a commercial kit (VYBRANT™ MTT Cell Proliferation Assay Kit,Catalogue no. V13154) from Thermo Fisher Scientific. The colon cancercells were seeded at a density of 10⁴ cells/well in 96-well platescontaining DMEM medium supplemented with 10% fetal bovine serum and 1%antibiotic mixture (Penicillin-Streptomycin). Different amounts of zincoxide nanoflowers and nanospheres were added to produce finalconcentrations of 1 mg/mL, 0.4 mg/mL, and 0.2 mg/mL within the wells.The control samples have neither nanoflowers nor nanospheres added. Thecells were maintained in a humidified atmosphere with 5% CO₂ at 37° C.and were incubated for 24 h. After incubation, the culture medium wasremoved and the wells were washed twice with phosphate-buffered saline(PBS). After adding fresh medium and 10 μL of MTT solution to each well,the cells were further incubated for 4 h. After incubation, the mediumwas removed, and 50 μL sterile DMSO was added to each well. Theabsorbance of each well was recorded on a SYNERGY Neo2 multi-modemicroplate reader (Biotek) at 570 nm. The cell viability was thencalculated using the following formula:

Cell viability (%)=absorbance of sample/absorbance of control×100%.

The cell viabilities observed for different amounts of nanospheres andnanoflowers are shown in FIG. 12. This graph shows that higherconcentrations of ZnO nanospheres or ZnO nanoflowers lead to lower cellviabilities, which is also an indicator of greater growth inhibition.

The procedures as described above involved different methods for thesynthesis of zinc oxide (ZnO) nanoparticles with different morphologies(nanoflowers and nanospheres) and sizes using zinc nitrate hexahydrateas a precursor. The prepared zinc oxide nanoparticles were characterizedby X-ray powder diffraction (XRD), scanning electron microscopy (SEM),UV-Vis diffuse reflectance spectrophotometry and BET surface areaanalysis. The potential application of zinc oxide nanoparticles wasevaluated for the (i) photocatalytic degradation of environmentalpollutant (Methyl orange), and (ii) biological application (cytotoxicityactivity). It was observed that both zinc oxide nanoflowers andnanospheres exhibited good photocatalytic degradation of methyl orange.Cytotoxicity of zinc oxide nanoflowers and nanospheres was evaluatedagainst human colon cancer cell line HCT116 by MTT assay. Both zincoxide nanoflowers and nanospheres suppressed growth of cancer cells andgrowth inhibition was 56.62-72.86% and 57.04-75.65% respectively over arange of concentrations (0.2-1 mg/mL).

1. A method for treating a colon cancer in a mammal, comprising:administering, to the mammal, a therapeutically effective dose of ZnOparticles having substantially spherical shapes with diameters of 220nm-3.5 μm and BET surface areas of 8-25 m²/g.
 2. The method of claim 1,wherein the therapeutically effective dose is 0.1 to 5 g of ZnOparticles per kg of the mammal per day.
 3. The method of claim 1,wherein the diameters are 1.5-3.5 μm, and the BET surface areas are15-25 m²/g.
 4. The method of claim 1, wherein the ZnO particles areporous with pore sizes of 20-35 nm.
 5. The method of claim 1, wherein asurface of the ZnO particles has nanopetals of 50-200 nm thicknessextending 100-500 nm from the surface, the nanopetals traversing thesurface with lengths of 500 nm-2 μm.
 6. The method of claim 1, whereinthe ZnO particles are administered as a part of a composition, whereinthe composition further comprises a food product, a pharmaceuticallyacceptable excipient, a pharmaceutically acceptable carrier, or anantioxidant.
 7. The method of claim 1, wherein the mammal is a human, anon-human primate, a dog, a cat, a horse, a cow, a goat, a rat, a pig, arabbit, or a mouse.
 8. The method of claim 1, wherein the ZnO particlesconsist essentially of ZnO.
 9. The method of claim 1, wherein the ZnOparticles contact a first population of colon cancer cells in the coloncancer, wherein 24 hours after the contacting, the first population hasa growth inhibition of 60-80% in relation to a second population ofcolon cancer cells in the colon cancer that were not contacted.
 10. Amethod of reducing a concentration of an organic contaminant in anaqueous solution, the method comprising: contacting ZnO nanoflowers withthe aqueous solution comprising the contaminant at a contaminantconcentration of 1 mg/L-1 g/L; and irradiating the ZnO nanoflowers whilein contact with the aqueous solution; wherein the ZnO nanoflowers have agenerally spherical shape with a diameter of 1.5-3.5 μm, wherein asurface of the ZnO nanoflowers has nanopetals of 50-200 nm thicknessextending 100-500 nm from the surface, the nanopetals traversing thesurface with lengths of 500 nm-2 μm, and wherein the ZnO nanoflowersreduce the concentration of the organic contaminant in the aqueoussolution by adsorption and/or photocatalytic degradation.
 11. The methodof claim 10, wherein the ZnO nanoflowers are dispersed within theaqueous solution at a concentration of 0.5-100 mg/L.
 12. The method ofclaim 10, wherein the irradiating is 15-30 minutes of UV lightirradiation, and wherein the concentration of the organic contaminantafter the irradiating is 40-50% of the concentration of the organiccontaminant before the irradiating.
 13. The method of claim 10, whereinthe organic contaminant is at least one selected from the groupconsisting of pharmaceutical compound, a dye, a metabolite, a microbialtoxin, an herbicide, a pesticide, and a steroid.
 14. The method of claim10, wherein the ZnO nanoflowers are made by heating an aqueous Zn²⁺solution with sodium hydroxide.
 15. The method of claim 14, wherein theaqueous Zn²⁺ solution comprises Zn(NO₃)₂.
 16. The method of claim 10,wherein the ZnO nanoflowers consist essentially of ZnO.
 17. The methodof claim 10, wherein the ZnO nanoflowers have a band gap energy of2.92-3.31 eV.
 18. The method of claim 10, wherein the irradiating usessunlight as an irradiation source.
 19. The method of claim 10, furthercomprising contacting the aqueous solution with a second absorbent or asecond photocatalyst to reduce the concentration of the contaminantand/or reduce a concentration of a second contaminant in the aqueoussolution.
 20. The method of claim 10, further comprising: rinsing theZnO nanoflowers to produce cleaned ZnO nanoflowers; and reusing thecleaned ZnO nanoflowers, which maintain a contaminant reduction capacityfor at least 5 purification cycles.